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At least 19 recordsLinked to original sources

Effects of poly[d(pGpT).d(pApC)] and poly[d(pCpG).d(pCpG)] repeats on homologous recombination in somatic cells.

Sequencing studies have shown that in somatic cells alternating runs of purines and pyrimidines are frequently associated with recombination crossover points. To test whether such sequences actually promote recombination, we have examined the effects of poly[d(pGpT).d(pApC)] and poly[d(pCpG).d(pCpG)] repeats on a homologous recombination event. The parental molecule used in this study, pSVLD, is capable of generating wild-type simian virus 40 DNA via recombination across two 751-base-pair regions of homology and has been described previously (Miller et al., Proc. Natl. Acad. Sci. USA 81:7534-7538, 1984). Single inserts of either a poly[d(pGpT).d(pApC)] repeat or a poly[d(pCpG).d(pCpG)] repeat were positioned adjacent to one region of homology in such a way that the recombination product, wild-type simian virus 40 DNA, could be formed only by recombination within the homologies and not by recombination across the alternating purine-pyrimidine repeats. We have found that upon transfection of test DNAs into simian cells, a poly[d(pCpG).d(pCpG)] repeat enhanced homologous recombination 10- to 15-fold, whereas a poly[d(pGpT).d(pApC)] repeat had less effect. These results are discussed in terms of the features of these repeats that might be responsible for promoting homologous recombination.

Animals↗

[The use of phase diagrams in the crystallization of oligonucleotide duplexes. I. A model of crystallization of the (pGpT)n.(pApC)n + spermine system].

A set of experimental phase diagrams revealing the region of existence of microcrystals in mixture "(pGpT)n.(pApC)n+spermine", n = 2,3,4, was obtained. All diagrams are wedge-like with the slope of the upper branch and the level of the lower one depending on the oligonucleotidd length. The presence of MPD, MgCl2 and NaCl changes the form of the diagrams in a different manner. A model explaining the peculiar features of the diagrams for mixture "oligonucleotide duplex+spermine" is proposed. The analysis of the diagrams was carried out on the basis of this model and the values of the binding constants for binding of spermine and Mg2+ to duplexes were estimated. Some conclusions about the types of complexes, which may form microcrystals in different regions of diagrams were made.

Binding Sites↗

[Construction of recombinant lentivirus vaccine with single round replication].

OBJECTIVE: To develop a safe and effective lentivirus vaccine model and provide insights into the development of other lentivirus vaccines. METHODS: In this study, a construct of pGPT was made by deleting env gene in the infectious Equine infectious anemia virus (EIAV) molecular clone of WU57. Since the overlaping of EIAV Rev gene with env gene, there was no Rev gene in the construct of pGPT. For compensation of Rev function, the construct of pGPTC was made by inserting 4 copies of constitutive RNA transport elements (CTEs) from Mason-Pfizer monkey virus into the construct of pGPT. In addition, a construct designated pTEB expressing EIAV Env protein was made while env gene-minus viruses were made by co-transfection of pGPT/pTEB or pGPTC/pTEB into 293 cells. Western blot was used to identify the development of recombinant virus particles. Then immunofluorescence assay was used to evaluate the infectivity of recombinant virus particles in vitro. RESULTS: EIAV proteins expression was detected in the supernatant of transfected 293 cells by Western blot within pGPTC/pTEB transfected cells. However, no evidence of EIAV proteins expression was observed within pGPT/pTEB transfected cells. EIAV proteins expression was detected in the first round but not in the second round infected EK cells with EIAV(GPTC) by immunofluorescence assay. CONCLUSION: Rev/RRE was necessary for expression of viral structural proteins; CTEs from Mason-Pfizer monkey virus was functionally interchangeable with EIAV Rev/RRE to help RNAs transportation out of nucleus to express structural proteins and EIAV particles were produced in the transfected 293 cells. A live EIAV recombinant virus with single round infection had been developed.

Animals↗

Whole-genome sequencing and characterization of Pseudomonas stutzeri P1 endophyte isolated from potato unveils plant growth-promoting and other traits.

Endophytic bacteria play an important role in plant growth promotion and stress tolerance, offering sustainable alternatives to chemical inputs in agriculture. In this study, an endophytic bacterial strain P1 was isolated and identified as Pseudomonas stutzeri, a plant-associated bacterium exhibiting multiple plant growth-promoting traits (PGPTs). Biochemical (qualitative and quantitative) and in vitro analyses demonstrated nitrogen fixation, phosphate solubilization, ammonia production, indole-3-acetic acid (IAA) production, biofilm formation, and tolerance to abiotic stresses, including salinity and drought. Furthermore, the P1 strain displayed strong biocontrol activity against the fungal pathogen Fusarium oxysporum f. sp. cumini, indicating its potential to mitigate biotic stress. Whole-genome sequencing generated a high-quality complete genome of 4,758,235 bp. Functional annotation showed enrichment of metabolic pathways associated with plant-microbe interactions and environmental adaptation. Further analyses using KEGG and PGPT-pred data confirmed the presence of genes associated with direct and indirect PGPT, such as nitrogen fixation, phosphate solubilization, biofilm formation, and stress tolerance. The genome also contained genes related to CAZymes, adhesion, and motility, highlighting a strong plant association, whereas the genome lacked major virulence factors and antimicrobial traits, supporting the non-pathogenic nature of the P1 strain. Overall, these findings demonstrate the potential of P1 as a promising bioinoculant candidate for sustainable agriculture in the potato sector.

PGPT-associated genes↗

Mercuric chloride-induced physiological dysfunction in Notopterus notopterus.

The freshwater fish Notopterus notopterus were exposed to various sublethal concentrations (0.088, 0.044, 0.029, 0.022, and 0.017) of mercuric chloride (HgCl2) mg/liter for 15, 30, 45, and 60 days, and their effects on glucose, lactate, cholesterol, plasma glutamic oxalacetic transaminase (PGOT) and plasma glutamic pyruvic transaminase (PGPT) of the blood were observed. Lactate increased significantly (P less than 0.05; P less than 0.01; P less than 0.001) and glucose. PGOT, and PGPT increased significantly (P less than 0.05; P less than 0.01). However, cholesterol decreased significantly (P less than 0.05; P less than 0.01; P less than 0.001) in the blood of N. notopterus.

Alanine Transaminase↗

Integrated functional genomics and safety assessment of plant-growth-promoting Caryophanales from post-maize-cultivation soils.

This study aimed to evaluate six environmental bacterial strains isolated from post-maize cultivation soils as candidates for agricultural biopreparation development, using an integrated functional genomic and safety assessment framework. Building on experimental validation of plant-growth-promoting activities, the analysis included: plant-growth-promoting traits (PGPT-Pred) using PLABase; carbohydrate-active enzymes (CAZymes) relevant for lignocellulosic crop residue degradation (dbCAN3); secondary metabolite profiles (antiSMASH); and screening for virulence factors and antibiotic resistance genes (ABRicate, BTyper3).All analyzed strains possess 1,449-1,617 predicted PGPT-encoding genes (24.1-35.9% of total genes), which are strongly shaped by taxonomic relatedness, as confirmed by congruence testing against ANI-based genomic divergence. Paenibacillus amylolyticus 5mez and Priestia megaterium 7psych showed distinct functional profiles compared to Bacillus spp., while Bacillus subtilis sensu lato strains were most similar to each other. Genomic predictions suggest involvement in nutrient acquisition (N, P, K, Fe) and stress mitigation. Secondary metabolite analysis revealed high biosynthetic potential, with non-Bacillus species harbouring a large proportion of unknown gene clusters, indicating underexplored metabolite diversity. CAZyme profiling identified P. amylolyticus 5mez as the most enzyme-rich strain, while B. cereus s.s. zielonkawy showed ligninolytic potential despite low overall CAZyme abundance. The safety assessment identified B. cereus s.s. zielonkawy as toxigenic and unsuitable for use. Of the remaining strains, P. amylolyticus 5mez and Pr. megaterium 7psych demonstrated the most favourable safety profiles, exhibiting no detectable virulence factors or antibiotic resistance genes, justifying their priority use in agricultural biopreparations, pending phenotypic validation. Given the high-dimensional, low-sample-size nature of multi-trait datasets in applied microbial genomics, tailored statistical approaches, including noise-reduction-validated PCA and distance-based congruence testing, were applied; their rationale and limitations are discussed.

Soil Microbiology↗

Potentiation of carbon tetrachloride hepatotoxicity by piperine.

The effect of piperine on CCl4-induced hepatotoxicity was investigated in rats. Piperine pretreatment potentiated the hepatotoxicity of CCl4 in a dose-dependent manner. The maximum potentiation occurred when piperine at a dose of 100 mg/kg BW was intragastrically administered 4 h prior to an intraperitoneal injection of CCl4, at which time the activities of plasma glutamic pyruvic transaminase (PGPT) and plasma glutamic oxaloacetic transaminase (PGOT) were elevated by 70-80%. Concurrent with the rise in PGPT and PGOT activities, the accumulation of hepatic triglyceride increased whereas the plasma level of triglyceride decreased. Piperine pretreatment also potentiated CCl4-induced lipid peroxidation in the liver. The extent of potentiation correlated well with the rise of hepatic enzyme activity in plasma. In the in vitro system in which the tissue was preincubated with piperine and CCl4 was added into the incubation medium, piperine also exhibited a concentration dependent potentiation on CCl4-induced lipid peroxidation and on the activity of NADPH-cytochrome c-reductase. The results indicated that piperine potentiated CCl4-induced hepatotoxicity by interacting with liver cells and increased the activity of NADPH-cytochrome c reductase. The increase in activity of this enzyme accelerated biotransformation of CCl4, thereby increasing lipid peroxidation and enhancing hepatotoxicity.

Alanine Transaminase↗

[The length of DNA determines the degree of regularity of crystals of the cro-repressor complex].

The DNA-cro-repressor complex crystals have been obtained, five DNA fragments of the same nucleotide sequence and different length being used. The rotation function for crystals of complexes with hexamer (pGpT)3 . (pApC)3 and with octamer (pGpT)3 . (pApC)3 have been calculated. The order of cro-DNA complex crystals is shown to vary with DNA length, the crystal of the complex with octamer being the most perfect among all investigated complexes.

Base Sequence↗

A conserved sequence motif at the 5' terminus of the Southampton virus genome is characteristic of the Caliciviridae.

We have determined the 5'terminal cDNA sequence for the genome of Southampton virus, a recently characterized, human, small round-structured virus (SRSV). Genomic RNA was extracted directly from a stool sample and amplified by RT-PCR by homopolymer tailing of the 3' terminus of the cDNA. The additional sequence increases the overall length of the Southampton virus genome by 12 nucleotides, resulting in a significant change to the genome organization by extending the first large open reading frame (ORF) by 51 amino acids. The 5'terminal bases pGpT and the presence of conserved genome and putative subgenomic RNA terminal motifs are now prominent features shared between the human SRSV Southampton virus and the animal caliciviruses rabbit hemorrhagic disease virus and feline calicivirus.

Amino Acid Sequence↗

Nuclease P1-mediated fluorescence postlabeling assay of AAF modified DNA model d(TACGTA) and calf-thymus DNA.

Fluorescence postlabeling assay for DNA damage combines enzymatic digestion of modified DNA to nucleoside monophosphates and fluorescence postlabeling. However, to facilitate the quantitative release of bulky adducts that are not readily obtained as mononucleotides, a different mechanism is essential. In order to test the application potential of nuclease P1-mediated fluorescence postlabeling to assay bulky adduct, d(TACGTA) was reacted with N-acetoxy-2-acetylamino-fluorene. The major product, characterized by nmr as AAF adduct of the guanine moiety at the C-8 position, was used as a DNA model. Nuclease P1 digestion of the modified oligomer excised the adduct in two forms (3:1) which were identified as AAF modified d(pGpT) and dpG respectively by cochromotography with authentic markers. Fluorescence postlabeling assay of AAF modified d(TACGTA) detected both forms of the excised adduct. The application of the overall procedure to assay AAF modified calf-thymus DNA demonstrated that the extension of fluorescence postlabeling technique from the mononucleotide to the dinucleotide version expands the scope of the assay.

Acetoxyacetylaminofluorene↗

Effects of cortisol pretreatment on the acute hepatotoxicity of aflatoxin B1.

Effects of cortisol pretreatment on acute hepatotoxicity induced by aflatoxin B1 (AFB1) were investigated in female rats. Pretreatment of cortisol (1.0-10.0 mg/kg body weight) for 7 consecutive days markedly increased mortality rate, activity of plasma glutamic pyruvic transaminase (PGPT), plasma glutamic oxaloacetic transaminase (PGOT) and liver triglycerides induced by AFB1 (3.0 mg/kg body weight). The potentiating action of cortisol on hepatic necrosis of AFB1 showed a dose-dependent pattern. The possible mechanism of its action may be related to an increase in the activity of aniline hydroxylase and formation of AFB1-2,3-epoxide which in turn caused a marked increase in AFB1 binding to hepatic DNA and proteins, and lipid peroxide formation. Therefore, the potentiating action of cortisol on AFB1 hepatotoxicity may possibly increase the damage to DNA and membranes of various organelles.

Aflatoxin B1↗

Time-course effects of ethanol pretreatment on hepatic necrosis and fat accumulation induced by aflatoxin B1 in the rat.

Effect of ethanol pretreatment on acute hepatotoxicity and hepatic fat accumulation induced by aflatoxin B1 (AFB1) was followed up to 120 h in male Wistar rats. Pretreatment with 4 oral doses of ethanol (4.0 g/kg body wt. each) at 48, 45, 24 and 21 h prior to AFB1 (2.0 mg/kg body wt.) single intraperitoneal administration caused a significant increase in the activity of plasma glutamic oxaloacetic transaminase (PGOT, 2.4-fold), plasma glutamic pyruvic transaminase (PGPT, 2.8-fold), liver triglycerides (2.3-fold) and the severity of liver necrosis at 72 h after AFB1 administration. The effect of ethanol pretreatment on an increase in the accumulation of liver cholesterol and cholesterol esters induced by AFB1 is additive in nature. In a time-course study, it was shown that liver necrosis and triglyceride, cholesterol and cholesterol ester accumulation occurred simultaneously in both groups of rats treated with AFB1 and ethanol-AFB1. These results suggest that fat accumulation per se is not a primary cause of liver necrosis induced by AFB1 and ethanol-AFB1.

Adipose Tissue, Brown↗

Kinetic studies of drug-dinucleotide complexes.

Three classes of kinetic behavior are observed in the complexes of actinomycin or ethidium with deoxydinucleotides. First, the initial dinucleotide binding to form a 1:1 complex is a rapid bimolecular process, whose rate could be measured for combination of actinomycin with d(pTpG) d(pGpT), d(pGpA), d(pGpG) d(pCpGpG), and d(pCpG) andfor combination of ethidium with d(pGpC). Second, with one exception, all reactions in which a second dinucleotide is added to form a 2:1 dinucleotide-drug complex are limited by a first-order step at high concentration. This class includes the combination of actinomycin with all dinucleotides tested except d(pGpC), and the reaction of ethidium with nucleotides of complementary sequence pyrimidine-purine, such as d(pCpG). The final class is the special case of d(pGpC) interacting to form a 2:1 complex with actinomycin. Third-order kinetics is observed, with no evidence for a first-order, rate-limiting step.

Chemical Phenomena↗

Phase diagrams for DNA crystallization systems.

Phase diagrams for several oligonucleotide duplex-spermine systems have been constructed. These diagrams characterize the duplex and spermine concentrations ranges in which crystalline precipitates are formed. All of them are wedge-like form. The slope of the upper branch of the diagram is determined by the oligonucleotide length. The position of the lower branch depends on both the nucleotide sequence and its length. The position of the lower branch depends on both the nucleotide sequence and its length. It has been shown that the addition to the system of MgCl2 and NaCl salts and MPD results in specific changes in the diagrams. A model for oligonucleotide duplex-spermine system has been suggested which explains the main characteristic features of the obtained phase diagrams. The experimental phase diagrams for the (pGpT)n (pApC)n-spermine system (n = 2,3,4) have been analyzed ion terms of this model and the values of the binding constants of spermine and Mg2+ ions binding to duplexes have been determined. It permitted to identify the complexes that precipitated in different regions of the phase diagrams under various conditions. The diagram obtained in the presence of a cobalt hexammine counterion is also considered. It has been shown that this phase diagram, in general, is similar to those obtained for the oligonucleotide duplex-spermine system.

Chemical Phenomena↗

Penicillopepsin-JT2, a recombinant enzyme from Penicillium janthinellum and the contribution of a hydrogen bond in subsite S3 to k(cat).

The nucleotide sequence of the gene (pepA) of a zymogen of an aspartic proteinase from Penicillium janthinellum with a 71% identity in the deduced amino acid sequence to penicillopepsin (which we propose to call penicillopepsin-JT1) has been determined. The gene consists of 60 codons for a putative leader sequence of 20 amino acid residues, a sequence of about 150 nucleotides that probably codes for an activation peptide and a sequence with two introns that codes for the active aspartic proteinase. This gene, inserted into the expression vector pGPT-pyrG1, was expressed in an aspartic proteinase-free strain of Aspergillus niger var. awamori in high yield as a glycosylated form of the active enzyme that we call penicillopepsin-JT2. After removal of the carbohydrate component with endoglycosidase H, its relative molecular mass is between 33,700 and 34,000. Its kinetic properties, especially the rate-enhancing effects of the presence of alanine residues in positions P3 and P2' of substrates, are similar to those of penicillopepsin-JT1, endothiapepsin, rhizopuspepsin, and pig pepsin. Earlier findings suggested that this rate-enhancing effect was due to a hydrogen bond between the -NH- of P3 and the hydrogen bond accepting oxygen of the side chain of the fourth amino acid residue C-terminal to Asp215. Thr219 of penicillopepsin-JT2 was mutated to Ser, Val, Gly, and Ala. Thr219Ser showed an increase in k(cat) when a P3 residue was present in the substrate, which was similar to that of the wild-type, whereas the mutants Thr219Val, Thr219Gly, and Thr219Ala showed no significant increase when a P3 residue was added. The results show that the putative hydrogen bond alone is responsible for the increase. We propose that by locking the -NH- of P3 to the enzyme, the scissile peptide bond between P1 and P1' becomes distorted toward a tetrahedral conformation and becomes more susceptible to nucleophilic attack by the catalytic apparatus without the need of a conformational change in the enzyme.

Amino Acid Sequence↗

Potentiation of aflatoxin B1 induced hepatotoxicity in male Wistar rats with ethanol pretreatment.

The interaction of ethanol and aflatoxin B1 (AFB1)-induced hepatotoxicity was studied in male Wistar rats using the activity of plasma GOT and GPT, liver triglyceride and histopathologic changes of liver necrosis as indices. Pretreatment of four oral doses of ethanol (4.0 g/kg BW each) at 48, 45, 24 and 21 hrs prior to AFB1 (0.5 to 2.0 mg/kg BW) single i.p. administration caused a significant increase in the activity of PGOT (6 folds) and PGPT (5 folds), liver triglycerides (2 folds) and severity of liver necrosis at 48 hrs after AFB1 administration. Ethanol pretreatment potentiated AFB1-induced hepatotoxicity by increasing MFO enzymes, aniline hydroxylase and p-nitroanisole-O-demethylase activity and lipid peroxidation, and decreasing in cytochrome b5, epoxide hydrolase activity and hepatic glutathione content. However, it did not cause any significant change in the activity of NADPH-cytochrome c reductase and glutathione-S-transferase and cytochrome P-450. These results suggest that potentiation of ethanol pretreatment on AFB1-induced hepatotoxicity may be due to an increase in the metabolic formation of AFB1-2, 3-oxide and subsequent binding to DNA.

Aflatoxin B1↗

Potentiation of carbon tetrachloride induced hepatotoxicity by thinner inhalation.

The interaction of thinner and carbon tetrachloride (CCl4) induced hepatotoxicity was studied in the rats using the activity of plasma GOT and GPT, liver triglyceride and histopathologic changes of liver necrosis as indices. The animals were housed in a chamber with the continuous flow of thinner vapour (1.11 g/litre/hr) for 2 hrs prior to i.p. administration of CCl4 (0.1 ml/kg BW) at 18 hrs after thinner inhalation. Thinner inhalation potentiated CCl4 induced hepatotoxicity in a dose-dependent manner. The maximal enhanced effect was observed at 24 hrs after CCl4 administration by which the activities of PGOT and PGPT were significantly increased (3 folds). Thinner itself caused an additive effect on CCl4 induced liver triglyceride accumulation. At 18 hrs after thinner inhalation, the activity of NADPH cytochrome C reductase was markedly increased (2.2 folds) but no change in the activity of aminopyrine N-demethylase which was able to increase the 14.CCl3 free radicals and binding to both the hepatic microsomal proteins (1.8 folds) and lipids (1.4 folds). In addition, thinner pretreatment somehow increased hepatic lipid peroxidation by 1.4 folds. These results suggest that thinner pretreatment causes an increase in mixed function oxidases to activate the formation of .CCl3 free radicals and binding to the microsomal proteins and lipids, which in turn stimulate hepatic damage via lipid peroxidation in the membrane.

Administration, Inhalation↗

[Use of phase diagrams in crystallization of oligonucleotide duplexes. II. Setting of the crystallized samples].

Oligonucleotide crystallization technique based on the method of phase diagrams is described in detail with (pGpT)3.(pApC)3 hexamer as an example. The key point of the technique consists of dividing the multiparameter crystallization space into a set of regions, each of which corresponds to the precipitation of a duplex in complex with a certain number of counterions.

Crystallization↗